The function and mechanism of AlkB Research Paper

The function and mechanism of AlkB - Research Paper Example Iron is known to interfere in the assays due to instability of AlkB/Fe+2 complexes in aerobic conditions; a problem that is overcome by replacing iron with other metals or performing assays under anaerobic conditions. The presence of iron bound to 2-oxoglutarate in the core of the enzyme has been established through over expression and isolation of native protein. Like all other Fe(II)/2-oxoglutarate-dependent dioxygenase superfamily enzymes AlkB too has a metal center exhibiting a UV-Vis band range of 52-580nm; that in absence of DNA is a five coordinate Fe2 center and changes to six coordinate center in presence of single stranded DNA. Like other members of the superfamily, AlkB too has a core jelly roll fold that is formed of 8 beta strands at the carboxy-terminus. The catalytic domain is contained within the carboxy-terminus; however additional features for substrate specificity lie outside the catalytic domain and the jelly roll fold. At the N-terminus additional beta strands an d alpha helices form a support scaffold for the catalytic domain and also the outer walls of Binding groove for DNA/RNA. In addition to these and many other structural similarities of AlkB to other members of Fe(II)/2-oxoglutarate-dependent dioxygenase superfamily; certain characteristics unique to AlkB include nucleotide binding lid, flipping mechanism motif. A common double stranded beta helix (DSBH) fold comprising of a large and a small beta sheet with iron core in between has the enzymes’ conserved residues. The first of the two distinct regions of DSBH includes a highly conserved iron binding region H131XD133XH187, wherein iron is bound to the 2-oxoglutarate in a bidentate form. The DSBH present the substrate binding site interacting exclusively with the damaged DNA/RNA strand through 2 amino terminal alpha helices and beta sheet loops that form a secondary structure called ‘lid’ (nucleotide recognition lid) over the active site. The flexible conformation o f the lid allows it its amino acids (Thr51, Tyr76 and Arg161) to bind to varied alkyl groups on its nucleotide substrates through H-bonding to phosphate group in the nucleotide backbone. As a consequence of this interaction the catalytic core of the AlkB, the enzyme loses its flexibility; undergoes a conformational change that disallows oxygen to reach the active site thereby preventing the oxidation of iron. It can thus be proposed that DNA binding if occurring after iron would lead to access of oxygen to active site. DNA/RNA repair mechanism of AlkB involves oxidative demethylation of nucleotides at the site of lesion which is accomplished through hydroxylation of methylated bases through oxidative decarboxylation of 2-oxoglutarate in the enzyme core. The latter as result is converted to succinate and CO2, and methyl group is released as formaldehyde. Though the actual mechanism is yet to be established, on the basis of studies on another enzyme of the same superfamily, TauD; the probable mechanism involving an oxidative intermediate to Trp178 has been proposed. The mechanism also verified through in vitro assays on purified AlkB involves the binding of Fe2 and 2-oxoglutarate to the enzyme core followed by binding of methylated middle base to the ‘lid’. This allows oxygen to reach the iron and form nucleophillic superoxo anion (OÂ ­2-) –Fe3. This then forms a bridged peroxo-type intermediate along with 2-oxoglutarate, that through decarboxylation of 2-oxoglutarate and cleavage of O2 forms Fe4-oxygen intermediate. Coupled to oxygen cleavage is formation of succinate and CO2 from 2-oxoglutarate. The intermediate on the other hand hydoxylates the methyl group at the nucleotide forming

The problem of sustainability at Marks and Spencer Essay

The problem of sustainability at Marks and Spencer - Essay Example The firm that is analyzed in the paper is Marks & Spencer, one of Europe’s leading retailers and a household name in the UK where it provides high-quality clothing, food, and home wares in about 30 countries through over 400 stores. It has thus employed many people worldwide and enjoys huge turnover. However, with the recent globalisation and increased competition, Marks & Spencer and other international companies face variant business challenges. The problem of sustaining the supply of high-quality clothing, food, and households is one of the major issues affecting Marks & Spencer. The core aim of Marks & Spencer is to provide great customer service since lack of it leads to customer disappointments and low turnover. Indeed, customers need full and consistent availability of high-quality services. In this case, Marks & Spencer aims at becoming the world's most sustainable major global retailer by 2015. The eight company executives who participated in the survey are specialist s who came from inside and outside Marks & Spencer. From the survey, we can establish that in 2007, Marks & Spencer launched a sustainability plan. As such, there is a clear relationship between sustainability and business models in various companies. This reflects an alignment between sustainability priorities and organisations’ business priorities. Indeed, this link is very significant as it enables the management to address the issues of sustainability in relation to business operations since all business operations should take into account the aspect of sustainability to guarantee a going concern for the company.... ctually, the survey reveals that Marks  &Spencer  aligns the concept of sustainability with core strategy in all business operations with an aim of realising sustainability in the quickest time possible. Indeed, most respondents established a clear correlation between the business strategy of the company and sustainability. However, many respondents established that there are wide gaps between the leaders’ aspirations towards sustainability and how sustainability applies within their organisations. This gap hinders the implementation of the sustainability policy and the success thereof. At the same time, we can derive form the survey that sustainability in any company faces internal and external challenges (Hogevold & Svensson 2012, p.142-151). Indeed, at Marks & Spencer, the current economy denies them an avenue to foster sustainability and its importance. Moreover, we can establish from the survey that companies face the challenge of balancing between sustainability prin ciples and sustainability objectives on one side and the actions and investments on another side. This seeks to ensure that we get maximum sustainability returns from potential investments. In addition, we can ascertain that being dependent on the importance of sustainability to the future business endeavours, the management faces a challenge of aligning the organisation’s sustainability practices with the principles of the company. As such, the management faces specific challenges of recruiting effective employees and personnel with the necessary skills to pursue the sustainability objectives as well as the challenge of making tradeoffs in other business areas. At the same time, sustainability faces a challenge of securing employee buy-in to business model changes (Deloitte 2010, p. 2-9). Most